Transplantation represents the standard-of-care for the treatment of many diseases characterized by end-stage organ failure. After transplantation, patients must rigidly adhere to lifelong, multi-agent treatment regimens that dramatically increase the risks of cardiovascular disease, infections and malignancies. Immune tolerance, the phenomenon by which the allograft is accepted without immunosuppression while preserving the recipient's protective immunity, represents a solution to the problems of acute and chronic rejection and the resulting long-term reliance on toxic immunosuppressive therapies. The development of tolerogenic strategies could not only reduce the risk of these life-threatening complications, but also greatly expand the application of organ, tissue and cellular transplantation for diseases such as the hemoglobinopathies and genetic immunodeficiencies, Type I diabetes, and possibly other autoimmune diseases. In rodent models, successful solid-organ transplantation tolerance has been created through strategies coupling hematopoietic chimerism-induction with T cell costimulation blockade. However, given the significant differences between the rodent and human immune systems, these strategies require rigorous testing in a translational model prior to their clinical application. Rhesus macaque non-human primate models have a number of important attributes that allow them to serve as critical preclinical models in order to bridge the basic insights gained in mice to their application to patient care. In this project, we will take advantage of our ability to induce chimerism using mobilized peripheral blood stem cells from living Rhesus macaque donors to perform a systematic analysis of the impact that a costimulation blockade and chimerism-based tolerance induction strategy has on transplant pairs having varying degrees of MHC disparity. These studies will focus on the efficacy of the addition of adoptive immunotherapies to our standard chimerism-induction regimen in increasing chimerism stability and immune competence after transplant. The unifying purpose of our proposal is to develop clinically applicable protocols for the induction of tolerance to solid organ allografts while preserving immune competence in the transplant recipient. Specifically, in this proposal, we will determine 1) whether adoptive immunotherapy using regulatory T cells improves the stability of mixed chimerism and the induction of transplantation tolerance;and 2) whether adoptive immunotherapy using donor lymphocyte infusions improves immune competence after the induction of mixed hematopoietic chimerism across MHC barriers.